Project Summary BRAF V600 mutations occur in ~10% of colorectal cancers (CRCs), leading to constitutive activation of the MAPK signaling pathway, and confer a ~2-fold increase in mortality relative to BRAF wildtype CRC. Thus, novel effective therapies for this disease are critically needed. While RAF inhibitors (RAFi), such as vemurafenib and dabrafenib, are highly effective in BRAF mutant melanoma (~60-80% response rate), the response rate to RAFi monotherapy in BRAF mutant CRC is only ~5%. Our initial efforts to define the resistance mechanisms operant in BRAF mutant CRC have led to novel clinical trials of RAFi combinations. Through this focused integration of laboratory models, clinical trials, and analysis of clinical specimens, significant advances in the care of BRAF mutant CRC patients have been achieved, with response rates increasing from ~5% to ~40% in the last few years. Still, a substantial percent of patients fail to respond to therapy, and those patients that do respond eventually develop resistance. Accordingly, further optimization of therapy is critically needed for BRAF mutant CRC. Therefore, we propose an innovative, highly translational approach leveraging detailed signaling studies utilizing established and patient-derived tumor models, comprehensive molecular assessment and characterization of patient-derived tumor models from tumor biopsies from BRAFm CRC patients enrolled in cutting-edge clinical trials, and serial liquid biopsy analysis of plasma ctDNA to define primary and acquired resistance mechanisms and the role of heterogeneity in BRAFm CRC. We will also evaluate a novel convergent inhibition strategy employing ERK inhibitors, supported by our preliminary data, as a potential strategy to overcome resistance. This proposed work will provide key insights to guide development of novel and more effective therapeutic strategies for future clinical trials.